Dr. John Hempel

Genetics
K. Arndt
T.-L. Ashman
G. Campbell
D. Chapman
G. Hatfull
J. Hildebrand
L. Jacobson
S. Kalisz
J. Martens
W. Saunders
B. Stronach
S. Tonsor
R. Wood

Microbiology
J. Boyle
G. Hatfull
R. Hendrix
J. Lawrence
J. Pipas
M. Popa
R.L. Duda
S. Godfrey
V. Oke

Molecular Biology
K. Arndt
J. Franzen
P. Grabowski
G. Hatfull
R. Hendrix
L. Jen-Jacobson
J. Martens
C. Peebles
J. Pipas
J. Rosenberg
A. Schwacha
C. Walsh

Plant Biology
T.-L. Ashman
W. Carson
S. Kalisz
V. Oke
C. Partanen
S. Tonsor
B. Traw

Science Education
A. Bledsoe
K. Curto
L. Daniels
S. Godfrey
N. Kaufmann
C. LaFave
J. Newman
E. Polinko
M. Popa
L. Roberts
T. Seiflein
R. Sherwin
A. Slinskey Legg

Structural Biology
M. Grabe
J. Hempel
R. Hendrix
L. Jen-Jacobson
J. Rosenberg
A. VanDemark

Former Faculty

Photo of Dr.
Hempel

Structure/Function Relationships of Proteins

Research Associate Professor

Dr. Hempel received his Ph.D. in 1981 at Rutgers University and joined the Department in 1996.

Currently, Dr. Hempel is not accepting graduate students in his laboratory. Dr. Hempel is accepting undergraduate researchers, and does sponsor students in other laboratories.

Professional Interests - Publications - Contact Information - Lab Personnel

Professional Interests of John Hempel

Aldehyde dehydrogenases (ALDHs) mediate NAD-coupled oxidation of aldehydes to carboxylic acids. ALDH is probably most widely known for its role in clearance of acetaldehyde, from beverage alcohol, by conversion to acetic acid. However, the ALDH family is a highly divergent one. A variety different forms with clear evolutionary relationships are known in humans. Some have broad substrate specificities (ALDH1 and ALDH2) and are primarily found in the liver. For instance, no physiological role is known for ALDH2, yet a single amino acid substitution (E487K) is sufficient to abolish activity, even in heterozygotes, with no apparent adverse effect except that it provides the underlying cause of severe aversion to beverage alcohol in ca. 50% of Asian individuals. Interestingly, ALDH2 has recently also been identified as responsible for the initial step in the pathway leading to nitric oxide from nitroglycerin, as used in treating angina. (ALDH2 converts nitroglycerin to 1,2 glyceryldinitrate and nitrite.) Other ALDHs are tailored to specific reactions of intermediary metabolism (e.g. methyl malonic semialdehyde dehydrogenase in branched-chain amino acid metabolism).

ALDH3, which my lab focuses on, is a form widespread in non-hepatic tissues and which prefers aromatic aldehydes. It is also estimated to constitutes ~10% of all protein in corneal epithelial cells, perhaps as a defense mechanism against airborne aldehydes which could otherwise react with protein amino groups, forming Schiff bases. With reference to cancer, ALDH3 is also found in some hepatomas. It is of considerable relevance to the efficacy of cyclophosphamide, a pro-drug, which was recently described as "arguably the most successful anticancer agent ever synthesized". ALDH3 acts on aldophosphamide, the (aldehydic) immediate precursor of the active metabolite of cyclophosphamide, the phosphoramide mustard. Thus, increased ALDH3 activity tempers the (anti-tumor) efficacy of cyclophosphamide. ALDH3 also acts to neutralize toxic aldehyde products generated by peroxidation of membrane lipids, a process elevated in alcoholism and other pathologies, and which is a factor in the cornea as well. In addition, deficiency of the membrane-bound (microsomal) form of ALDH3, "fatty" aldehyde dehydrogenase, which acts on long-chain aldehydes, results in Sjögren-Larsson Syndrome, a rare but severe neurocutaneous disorder.

Clearly, with such activities as these, detailed knowledge of the molecular structure of any ALDH would be valuable. In collaboration together with colleagues at the Universities of Georgia and South Dakota, the structure of ALDH3 was solved, at 2.6 &ARing;nstroms resolution.

In addition, now with sequences known for over 550 members of the ALDH extended family, comparison of residue exchanges are of great value in understanding the structure/function relationships involved in maintaining catalytic activity and tailoring individual enzymes to their particular substrate specificity. Our collaboration with Hugh Nicholas at the Pittsburgh Supercomputing Center (PSC) resulted in an alignment of 145 ALDHs which is electronically available. Many of our continuing efforts are grounded in this alignment

Subsequent results from site-directed mutagenesis illuminated the molecular basis of the ability of ALDH3 to use both NAD and NADP as coenzyme. Another group of mutants probed roles of the only two strictly-conserved, non-glycine residues in over 400 ALDHs, a glutamic acid and phenylalanine residue. Further, an algorithm developed at the PSC has been applied to an expanded alignment identify residues without prejudice which are most specific to the ALDH3 and other ALDH families. One of the residues diagnostic of ALDH3 (Asp-247) seemed to represent a linchpin tethering elements of the catalytic center. A mutant of this aspartic acid in fatty ALDH has been found in a Sjögren-Larsson patient, supporting its importance despite a lack of conservation across the entire ALDH family.

Our current focus is on understanding the full chemical mechanism of catalysis. These results used in tandem with quantum mechanical/molecular mechanical calculations from Troy Wymore's efforts at the PSC have been used to great advantage to gain insight into the chemical mechanism. Most recently, these simulations pointed to the chemically reasonable yet baffling formation of an adduct between the catalytic thiol and nicotinamide C-4. Since this would represent a dead-end complex we reasoned that these results must have been due to an incorrect starting data set, but within months of this finding, crystallographic evidence for precisely that adduct was independently found (Tsybovsky et al. Biochemistry 46: 2917 (2007); these authors also demonstrated the reversibility of the adduct), greatly supporting the validity of our computational work. In our most recent publication we have addressed the question of what conditions favor (and conversely block) adduct formation.

Publication Archive
71 Citations
58 Abstracts
1 PDFs

Recent Publications of John Hempel

Wymore, T., D.W. Ii, and J. Hempel (2007) Mechanistic implications of the cysteine-nicotinamide adduct in aldehyde dehydrogenase based on quantum mechanical/molecular mechanical simulations. Biochemistry 46:9495-9506

Hempel, J., H.B. Nicholas, S.T. Brown, and T. Wymore (2007) Unexpected encounters in simulations of the ALDH mechanism. Pp 9-13 in Enzymology and Molecular Biology of Carbonyl Metabolism 13, Weiner, H., E. Maser, R. Lindahl, and B. Plapp, Ed. Purdue University Press, Lafayette, IN

Hempel, J., S. Stanley, J. Perozich, T. Wymore, and H.B., J.r. Nicholas (2006) Residue conservations in aldehyde dehydrogenase gene fusion products reemphasize functional interpretations. Pp 8-14 in Enzymology and Molecular Biology of Carbonyl Metabolism 12, Weiner, H., B. Plapp, R. Lindahl, and E. Maser, Ed. Purdue University Press, Lafayette, IN

Wymore, T., J. Hempel, S.S. Cho, A.D. .J.r. Mackerell, H.B. .J.r. Nicholas, and D.W. .2.n. Deerfield (2004) Molecular recognition of aldehydes by aldehyde dehydrogenase and mechanism of nucleophile activation. Proteins Struct. Func. Bioinf. 57:758-771

Hempel, J., J. Perozich, T. Wymore, and H.B. Nicholas (2003) An algorithm for identification and ranking of family-specific residues, applied to the ALDH3 family. Chem. Biol. Interact. 143:23-28

Wymore, T., D.W. Deerfield, M.J. Feidl, J. Hempel, and H.B. Nicholas (2003) Initial catalytic events in class 3 aldehyde dehydrogenase: MM and QM/MM simulations. Chem. Biol. Interact. 143:75-84

Lee, J.Y.-Y., J. Hempel, and J.-S. Deng (2002) Anti-adenosine deaminase antibodies in lupus erythematosus. Lupus 11:168-174

Hempel, J., R. Lindahl, J. Perozich, B. Wang, I. Kuo, and H. Nicholas (2001) Beyond the catalytic core of ALDH: a web of important residues begins to emerge. Chem. Biol. Interact 130:39-46

Perozich, J., I. Kuo, R. Lindahl, and J. Hempel (2001) Coenzyme specificity in aldehyde dehydrogenase. Chem. Biol. Interact 130:115-124

Wymore, T., H.B. Nicholas, and J. Hempel (2001) Molecular dynamics simulation of class 3 aldehyde dehydrogenase. Chem. Biol. Interact 130:201-207

Hempel, J. (2001) An Orientation to Edman chemistry. Pp 102-122 in Practical Methods in Advanced Protein Chemistry, Brown, W.E., and G.C. Howard, Ed. CRC Press, Boca Raton

Hempel, J., I. Kuo, J. Perozich, B.C. Wang, R. Lindahl, and H. Nicholas (2001) Aldehyde dehydrogenase: maintaining critical active site geometry at motif 8 in the class 3 enzyme. Eur. J. Biochem. 268:722-726

Perozich, J., I. Kuo, J.S. Boesch, B.C. Wang, R. Lindahl, and J. Hempel (2000) Shifting the NAD/NADP preference in class 3 aldehyde dehydrogenase. Eur. J. Biochem. 267:6197-6203

How to Contact John Hempel

US Mail
University of Pittsburgh
Department of Biological Sciences
A356 Langley Hall
4249 Fifth Avenue
Pittsburgh, PA 15260

Phone, FAX, Internet
Office : (412) 624-0161
Lab : (412) 624-0106
FAX : (412) 624-4759
Email : hempel+@pitt.edu
Web :

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